Stabilized and lyophilized radiopharmaceutical agents

ABSTRACT

A novel method is set out of preparation of radioactive diagnostic radiopharmaceutical in a stable, shippable, lyophilized form by an apparatus designed to rapidly flash freeze and dehydrate a radiopharmaceutical composition to minimize auto radiolysis. The method proposes rapid cooling and removal of ambient vapor, and then ultra cold removal when the potential of explosive liquid oxygen is eliminated. The radioactive diagnostic radiopharmaceutical requires no further cold or refrigerated storage, including with respect to shipping, subsequent to stabilization. The preferred composition can be reconstituted “on site” by the addition of a suitable diluent to bring the radiopharmaceutical complex into solution at a desired concentration.

This is a continuation-in-part of a pending U.S. utility applicationSer. No. 10/904,099 entitled Stabilized and LyophilizedRadiopharmaceutical Agents which is a continuation-in-part ofprovisional application No. 60/580,455 entitled Stabilized andLyophilized Radiopharmaceutical Agents filed on Jun. 17, 2004 and aprovisional application No. 60/608,060 of that name filed on Sep. 8,2004, and a provisional application No. 60/522,619 filed on Oct. 20,2004, and a continuation-in-part of U.S. application Ser. No. 11/611,862filed Dec. 16, 2006 and Ser. No. 11/570,852 filed Dec. 18, 2006 whichare US national stage entries of PCT/US2005/21847 which claim priorityfrom the above 2004 applications and other applications as more fullyset forth in PCT/US2005/21847.

FIELD OF THE INVENTION

The present invention relates to the method of preparation andstabilization of a diagnostic or therapeutic radiopharmaceutical useful,for example, in mammalian imaging and cancer detection, and resultingcomposition. In particular, the present invention relates to the novelmethod of preparation of radioactive diagnostic radiopharmaceutical in astable, shippable, lyophilized form by an apparatus designed to rapidlyflash freeze and dehydrate a radiopharmaceutical composition to minimizeauto radiolysis, the novelty centering on rapid cooling and removal ofambient vapor, and then ultra cold removal when the potential ofexplosive liquid oxygen is eliminated. The radioactive diagnosticradiopharmaceutical requires no further cold or refrigerated storage,including with respect to shipping, subsequent to stabilization. Thepreferred composition can be reconstituted “on site” by the addition ofa suitable diluent to bring the radiopharmaceutical complex intosolution at a desired concentration at the time of administration to thepatient in need of a therapeutic or diagnostic radiopharmaceutical.Heading

SUMMARY OF THE INVENTION

The present invention is directed to a stable radioactive diagnosticradiopharmaceutical composition that may be formed without stabilizationadditives and to a method of preparing such a composition. Stabilizationadditives may be added. Traditional techniques for freeze-drying(lyophilization) are subject to the lengthy crystal formation time ofwater. The composition is formed by avoiding that lengthy crystalformation time and the concurrent loss of diagnostic specificity due toautoradiolysis of the radiopharmaceutical. The length of traditionalfreeze-drying techniques and loss of diagnostic specificity due toautoradiolysis interfere with the technical accuracy necessary fornuclear medicine.

The novel technique of the inventors involves utilization of flashfreeze techniques along with increasing the cold-exposed surface areaand then rapidly decreasing the vapor pressure as well as super coldfreeze drying of the radiopharmaceutical composition, the combination ofwhich results in extremely rapid freeze-drying/lyophilization, enablinguse of higher concentrations of radionuclides in the small scale amountsused in radiopharmaceutical imaging without damaging the ligands. Theradiopharmaceutical composition can be reconstituted immediately priorto administration with confidence of little or no ligand damage, orlittle or no damage to the non-radioactive bonds and chemical structureof the composition.

The preferred composition results from forming a complex between a gammaemitting radionuclide and a ligand in a suitable solvent, generally anaqueous solution and then lyophilizing the solution by use of smallquantities in large surface area vessels at vacuum pressure inconjunction with rapid sub-zero cooling. The radioactive diagnosticradiopharmaceutical in this invention requires no further cold orrefrigerated storage, including with respect to shipping, subsequent tostabilization. The lyophilized radiopharmaceutical composition isshipped and stored and is often reconstituted “on site” by the additionof a suitable diluent to bring the radiopharmaceutical complex intosolution at the time of administration to the patient in need of atherapeutic or diagnostic radiopharmaceutical. The present inventionfurther is directed to stable radioactive diagnostic radiopharmaceuticalcompositions prepared by this method.

BACKGROUND OF THE INVENTION

With the invention of the Gamma Camera, and, just as importantly, withthe invention of better high-speed imaging machines, pharmaceuticalsubstances with radioactive “tags” have become extremely important inmedical imaging and treatment. The concept is that a compound, or justas often, a part of a compound, called a ligand, sometimes referred toas an “agent” or which bonds to some other substance, is designed totarget a particular area of a mammal's body or a particular type oftissue or molecule in that body. The compound, ligand or agent will bereferred to as a ligand for convenience sake. The mammal this is mostoften used on is the human body, and references in this invention to ahuman are equally applicable to any mammal, or for that matter to anyanimal or plant.

For instance, certain ligands tend to concentrate in heart muscletissue. The concept behind radiopharmaceutical imaging is to “tag” thatligand with a radioactive substance, i.e. radioactively mark a substanceto create an “imaging agent,” so that a health care provider can findout where the ligand exists or is concentrating. By administering theradioactively tagged ligand, and placing the patient in an imagingmachine, a health care provider can “look inside” a patient's body toassist in therapy or diagnosis. If a person has poor heart circulation,the radionuclide tagged ligand, such as Tc 99m TIBI, will not bewell-circulated to areas of the heart muscle which have compromisedblood flow, enabling evaluation of a person's “heart condition.”Importantly, the health care provider can often “look inside” withouthaving to actually cut open or invade the body (non-invasive technique),or can minimize bodily invasion. Obviously, the continued presence ofradioactive substances is not desirable, so substances are selected witha short “half-life.” The half-life is a time defined as the time inwhich the radioactive emission declines by one-half. The diminution ofradioactivity is referred to as radioactive decay. Between the bodywashing out the radiopharmaceutical substances used in conjunction withthis invention, and the use of substances with a short half-life, theamount of a patient's radioactive exposure is minimized.

Radioactive pharmaceuticals are in common use in imaging studies to aidin the diagnosis of a wide variety of illnesses including cardiac, renaland neoplastic diseases. These pharmaceuticals, known in the art as“imaging agents,” typically are based on a gamma-emitting radionuclideattached to a carrier molecule or “ligand.” Gamma-emitting radionuclidesare the radionuclides of choice for conducting diagnostic imagingstudies because, while gamma emitting radiation is detectable withappropriate imaging equipment, it is substantially less-ionizing thanbeta or alpha radiation. Thus, gamma emitting radiation causes minimaldamage to targeted or surrounding tissues.

Radioactive pharmaceuticals now are finding increased use as diagnosticagents for finding neoplastic disorders, especially tumors. Diagnosticradiopharmaceuticals generally incorporate a gamma emittingradionuclide, the radiation emission being useful in the detection ofcertain neoplastic disorders.

The radioactive marking or tagging is often done by complexing theradioactive substance inside a group of ligands, that is surrounding itby a complex of ligands, so that the desired chemical characteristicsare expressed toward the exterior of the complex with the tag shieldedby the outer complex and simply carried along as a marker. The entirecomplex with the radioactive element, also called a radionuclide,functions as a radioactive marker, and can be more generally referred toas a radiopharmaceutical.

The use of small quantities of drugs used for such activities isdesirable for cost reasons, and it is desirable to minimize the amountof radioactive substance used.

While the efficacy of radioactive diagnostic and therapeutic agents isestablished, it is also well known that the emitted radiation can causesubstantial chemical damage or destabilization to various components inradiopharmaceutical preparations, referred to as autoradiolysis. Emittedradiation causes the generation of free radicals in water solutions,which free radicals are generally peroxides and superoxides. Such freeradicals can precipitate proteins present in the preparations, and cancause chemical damage to other substances present in the preparations.Free radicals are molecules with unbonded electrons that often resultbecause the emissions from the radioactive element can damage moleculesby knocking apart water molecules forming hydroxyl radicals and hydrogenradicals, leaving an element or compound with a shell of chargedelectrons which seek to bond with other molecules and atoms anddestabilize or change those molecules and atoms. The degradation anddestabilization of proteins and other components caused by the radiationis especially problematic in aqueous preparations. Under the presentart, the radiolysis causes the aqueous stored ligand and radioactiveisotope bonded to the ligand to degenerate and destroys the complexwhich renders it useless for imaging because the biologicalcharacteristics that localize the complex to a tissue are gone. Thedegradation or destabilization lowers or destroys the effectiveness ofradiopharmaceutical preparations, and has posed a serious problem in theart. Wahl, et al, Journal of Nuclear Medicine, Vol 31, Issue 1 84-89,discuss the fact that freezing radiolabeled antibodies at −70 degrees C.stabilizes the molecule for an indefinite period but 80 to 90% of theimmunoreactivity is lost in as little as 24 hours when stored at 4degrees C.

If the ligands are permitted to reside with the radioactive elements foran extended period, particularly in an aqueous (water-based) solution,the radiolysis is increased. Thus, any process to reduce the compoundsto dried form has to be rapid and yield predictable result. Further, toavoid the higher concentrations and protect the ligands, presently theradiopharmaceutical solution is diluted, but that in itself only slowsthe drying time and complicates the problem and increases theunpredictability of the non-radioisotope portion of theradiopharmaceutical because of radiolysis. Heating theradiopharmaceutical in solution to accelerate the drying and removal ofwater has the undesirable effect of potentially damaging the ligandsince chemical activity normally increases upon heating or injection ofenergy and therefore the effects of radiolysis are also increased duringthis prolonged drying period with heating. Most proteins are badlydamaged upon heating. Certain ligands, such as isonitrile, simplyevaporate and disappear upon heating. Further, minimization of localizedheating at an atomic scale is important to preserve both the smallquantities needed and to yield a specific concentration of desiredproduct.

Wolfangel, U.S. Pat. No. 5,219,556, Jun. 15, 1993, entitled stabilizedtherapeutic radiopharmaceutical complexes, expressed his concern asfollows: “The isotopes which are most useable with this process aredetermined by practical considerations. Again, Tc-99m would be a poorcandidate for use since its six-hour half-life makes lyophilizationimpractical, as the lyophilization step itself generally takes about 24hours to perform.”

Facially, the '556 invention seemed to identify a useful process andresulting composition, but the lyophilization step in '556 invention, asthe application stated, took about 24 hours. The '556 invention stated:“The lyophilization is carried out by pre-freezing the product, and thensubjecting the frozen product to a high vacuum to effect essentiallycomplete removal of water through the process of sublimation. Theresultant pellet contains the complex in an anhydrous form whichgenerally can be stored indefinitely, with practical consideration beinggiven to the half-life of the radionuclide. The intended period ofstorage for radiopharmaceutical products is thus practically limited bythe half-life of the radionuclides. In the case of Re-186, for example,the desired period of storage would range from 7 to about 30 days. Thus,this pellet can be shipped to the end users of the product andreconstituted with a diluent at the time of administration to thepatient with very little effort on the part of the health careprofessional and/or nuclear pharmacist.”

Because the procedures in '556 did not rapidly lyophilize the product,and contemplated a 24 hour period for lyophilization, the claims of '556invention were necessarily limited to utilization of a “therapeuticamount of an alpha- or beta-emitting radionuclide.” Wolfangel hadobserved that compounds with a half-life of at least 12 hours arepreferred. By contrast, the use of Tc-99m, which also emits gamma rays,with a half-life of only six hours, or the use of other similarlyshort-lived radioisotopes, becomes impractical.

In a recent comprehensive text on the subject of lyophilization, apre-eminent authority in the field made the following observations:

“Lyophilization is a multistage operation in which quite obviously eachstep is critical. The main actors of this scenario are all well knownand should be under strict control to achieve a successful operation.

The product, . . .

The surrounding “medium” . . .

The equipment, . . .

The process, which has to be adapted to individual cases according tothe specific requirements and low-temperature behavior of the differentproducts under treatment.

The final conditioning and storage parameters of the finished product,which will vary not only from one substance to another one but inrelationship with its “expected therapeutic life” and marketingconditions (i.e., vaccines for remote tropical countries, internationalbiological standards, etc.). In other words a freeze-dryer is not aconventional balance; it does not perform in the same way with differentproducts. There is no universal recipe for a successful freeze-dryingoperation and the repetitive claim that “this material cannot befreeze-dried” has no meaning until each successive step of the processhas been duly challenged with the product in a systematic andprofessional way and not by the all-too-common “trial-and-error” game.

The freeze-drying cycle. It is now well established that a freeze-dryingoperation includes:

-   -   The ad hoc preparation of the material (solid, liquid, paste        emulsion) to be processed taking great care not to impede its        fundamental properties.    -   The freezing step during which the material is hardened by low        temperatures. During this very critical period, all fluids        present become solid bodies, either crystalline, amorphous, or        glass. Most often water gives rise to a complex ice network but        it might also be imbedded in glassy structures or remain more or        less firmly bound within the interstitial structures. Solutes do        concentrate and might finally crystallize out. At the same time,        the volumetric expansion of the system might induce powerful        mechanical stresses that combine with the osmotic shock given by        the increasing concentration of interstitial fluids.    -   The sublimation phase or primary drying will follow when the        frozen material, placed under vacuum, is progressively heated to        deliver enough energy for the ice to sublimate. During this very        critical period a correct balance has to be adjusted between        heat input (heat transfer) and water sublimation (mass transfer)        so that drying can proceed without inducing adverse reactions in        the frozen material such as back melting, puffing, or collapse.        A continuous and precise adjustment of the operating pressure is        then compulsory in order to link the heat input to the        “evaporative possibilities” of the frozen material.    -   The desorption phase or secondary drying starts when ice is        being distilled away and a higher vacuum allows the progressive        extraction of bound water at above zero temperatures. This again        is not an easy task since overdrying might be as bad as        underdrying. For each product, an appropriate residual moisture        has to be reached under given temperatures and pressures.    -   Final conditioning and storage begins with the extraction of the        product from the equipment. During this operation great care has        to be taken not to lose the refined qualities that have been        achieved during the preceding steps. Thus, for vials, stoppering        under vacuum or neutral gas within the chamber is of current        practice. For products in bulk or in ampoules, extraction might        be done in a tight gas chamber by remote operation. Water,        oxygen, light, and contaminants are all important threats and        must be monitored and controlled.        -   Ultimate storage has to be carried according to the specific            “sensitivities” of the products (at room temperature, +4 C            −20 C). Again uncontrolled exposures to water vapor, oxygen            (air), light, excess heat, or nonsterile environment are            major factors to be considered. This obviously includes the            composition and qualify of the container itself, i.e., glass            elastomers of the stoppers, plastic or organic membranes.    -   At the end, we find the reconstitution phase. This can be done        in many different ways with water, balanced salt solutions, or        solvents either to restore the concentration of the initial        product or to reach a more concentrated or diluted product. For        surgical grafts or wound dressing, special procedures might be        requested. It is also possible to use the product as such, in        its dry state, in a subsequent solvent extraction process when        very dilute biochemicals have to be isolated from a large        hydrated mass, as is the case for marine invertebrates.        [emphasis added as underlined material; italics in original]”

Rey, Louis and May, Joan C., editors, Freeze-Drying/Lyophilization ofPharmaceutical and Biological Products, pp. (Marcel Dekker, Inc., NewYork, Basel 1999 (Nat'l Library of Medicine Call no. WI DR893B v. 961999)).

Professor Rey states in his introduction that he and two others led thefirst conference in 1958 on cryobiology, including freeze-drying ofpharmaceuticals. Id. at 2.

Juxtaposing the most important underlined material from the aboveexcerpt from Professor Rey's commentary based on his life-longexperience, the following important principles are stated to be the artas of 1999:

-   -   a) “Lyophilization is a multistage operation in which, quite        obviously, each step is critical. The main actors of this        scenario are all well known and should be under strict control        to achieve a successful operation.”    -   b) In other words, a freeze-dryer is not a conventional balance;        it does not perform in the same way with different products.        There is no universal recipe for a successful freeze-drying        operation and the repetitive claim that “this material cannot be        freeze-dried” has no meaning until each successive step of the        process has been duly challenged with the product in a        systematic and professional way and not by the all-too-common        “trial-and-error” game.    -   c) The freeze-drying cycle. It is now well established that a        freeze-drying operation includes: . . .        -   1). preparation of the material        -   2) The freezing step        -   3) The sublimation phase or primary drying will follow when            the frozen material, placed under vacuum, is progressively            heated to deliver enough energy for the ice to sublimate.            During this very critical period a correct balance has to be            adjusted between heat input (heat transfer) and water            sublimation (mass transfer) so that drying can proceed            without inducing adverse reactions in the frozen material            such as back melting, puffing, or collapse. A continuous and            precise adjustment of the operating pressure is then            compulsory in order to link the heat input to the            “evaporative possibilities” of the frozen material.        -   4) The desorption phase or secondary drying starts when ice            is being distilled away and a higher vacuum allows the            progressive extraction of bound water at above zero            temperatures. This again is not an easy task since            overdrying might be as bad as underdrying. For each product,            an appropriate residual moisture has to be reached under            given temperatures and pressures.        -   5) Final conditioning and storage . . .        -   6) Ultimate storage . . . . . . reconstitution phase . . .            [emphasis added as underlined material; italics in            original]”

The current patent art corroborates Professor Rey's assertion of theneed for progressive heating, particularly the Wolfangel '556 art, andCorbo et al art, U.S. Pat. No. 6,024,938, Feb. 15, 2000. Wolfangel '556contains the heating step that Professor Rey asserts is well-settled,while the present invention omits that step while achieving a superiorart, and sets out a procedure that is not isolated to a particularproduct, but as useful for all radiopharmaceuticals. Further, thepresent invention presents an advantage of rapidity of process not seenin the prior art.

Notably, Wolfangel not only specifically also includes a heating step,but simultaneously and specifically states his invention is notapplicable to short-half-life radionuclides. Corbo '938 also containsthe heating step, as does DeRosch, U.S. Pat. No. 6,428,768, Aug. 6,2002, and all except Wolfangel take upwards of 24 hours. Thus, the laterdeveloping art is in fact moving to longer periods of time,notwithstanding the possible aspiration to a shorter time.

This invention thus defies the conventional wisdom by omitting theheating step, but lyophilizing, and dehydrating, and thereby stabilizinga radiopharmaceutical capable of storage at room temperature by adifferent technique, thereby achieving a superior result as demonstratedby the comparative experimental results discussed momentarily.

Wolfangel '556 proposed in his example 1 to first lyophilize certaincompounds, add the radionuclide complex, sparge with gas, seal the vialand then heat it. Unfortunately, the heating to 100 degree C. rendersthe procedure useless in conjunction with most proteins or peptides, andmany commonly used complexes. Further, the proposal was to use 1 ml ofsodium perrhenate Re-186 containing 1 mg of rhenium, with water added toproduce 3 ml. The quantities contemplated were substantial and exposedthe workers to substantial amounts of radiation. In example 3, it wasproposed that the complex be frozen to −30 degree C. or colder and thenapply a vacuum, but it was proposed to apply shelf heat at 6 degree perhour until a product temperature of 30 degree C. was reached, at whichtime the temperature would be held for two hours. That would require 12hours. The procedure suffered from the infirmity of not quickly removingwater and therefore not preventing radiolysis of the water and notpreventing the generation of free radicals which damage the complexes.The second example 2 followed the first, but used smaller quantities,and proposed heating. Example 3 proposed heating to 85 degree C. for 30minutes which would destroy most proteins and thereafter freezing andlyophilizing the sealed vials.

For diagnostic imaging purposes, radiopharmaceuticals based on acoordination complex comprised of a gamma-emitting radionuclide and achelate have been used to provide both negative and positive images ofbody organs, skeletal images and the like. The Tc-99m skeletal imagingagents are well-known examples of such complexes. One drawback to theuse of these radioactive complexes is that while they are administeredto the patient in the form of a solution, neither the complexes per senor the solutions prepared from them are overly stable. Consequently,the coordination complex and solution to be administered commonly areprepared “on site,” that is, they are prepared by a nuclear pharmacistor health care technician just prior to conducting the study. Thepreparation of appropriate radiopharmaceutical compositions iscomplicated by the fact that several steps may be involved, during eachof which the health care worker must be shielded from the radionuclide.

The preparation of stable radiopharmaceutical diagnostic agents, due tothe type of radioactivity, presents even greater problems. These agentstypically are based on a relatively energetic gamma emittingradionuclide complexed with a chelate. Frequently, theradionuclide/chelate complex is in turn bound to a carrier moleculewhich bears a site-specific receptor. Thus, it is known that a gammaemitting radionuclide attached to a tumor-specific antibody or antibodyfragment can destroy targeted neoplastic or otherwise diseased cells viaexposure to the emitted ionizing radiation. Bi-functional chelatesuseful for attaching a diagnostic radionuclide to a carrier moleculesuch as an antibody are known in the art. See e.g. Meares et al., Anal.Biochem. 142:68-78 (1984).

For most imaging and diagnostic applications of radiopharmaceuticalcomplexes of the types mentioned above, the nonradioactive portion(s) ofthe complex is prepared and stored until time for administration to thepatient, at which time the radioactive portion of the complex is addedto form the radiopharmaceutical of interest. For example, attempts toprepare radionuclide-antibody complexes have resulted in complexes whichmust be administered to the patient just after preparation because, as aresult of radiolysis, immunoreactivity may decrease considerably afteraddition of the radionuclide to the antibody. In Mather et al., J. Nucl.Med., 28:1034-1036 (1987), a technique for labeling monoclonalantibodies with large activities of radio iodine using the reagentN-bromosuccinimide is described. The authors suggest that the antibodieslabeled in this manner be administered to the patient immediately afterpreparation to avoid losses of immunoreactivity. Other examples of thepreparation of the nonradioactive portion of the complex followed byon-site addition of the radioactive portion are disclosed in U.S. Pat.No. 4,652,440 (1987). Further, in many situations, the radioactivecomponent of the complex must be generated and/or purified at the timethe radiopharmaceutical is prepared for administration to the patient.U.S. Pat. No. 4,778,672 (1988) describes, for example, a method forpurifying pertechnetate and perrhenate for use in a radiopharmaceutical.

According to Wolfangel '556, EP 250,966 (1988) describes a method forobtaining a sterile, purified, complexed radioactive perrhenate from amixture which includes, in addition to the ligand-complexed radioactiveperrhenate, uncomplexed ligand, uncomplexed perrhenate, rhenium dioxideand various other compounds. Specifically, the application teaches amethod for purifying a complex of rhenium-186 and 1-hydroxyethylidenediphosphonate (HEDP) chelate from a crude solution. Because of theinstability of the complex, purification of the rhenium-HEDP complex bya low pressure or gravity flow chromatographic procedure is required.The purification procedure involves the aseptic collection of severalfractions, followed by a determination of which fractions should becombined. After combining the appropriate fractions, the fractions aresterile-filtered and diluted prior to injection into the patient. Thepurified rhenium-HEDP complex should be injected into the patient withinone hour of preparation to avoid the possibility of degradation. Therhenium complex may have to be purified twice before use, causinginconvenience and greater possibilities for radiation exposure to thehealth-care technician.

While the lyophilization process has been applied to various types ofpharmaceutical preparations in the past, the notion of lyophilizingshort lived gamma emitting radiopharmaceutical preparations has not beenaddressed. In part, this is believed to be due to skepticism of thoseskilled in the art that such a procedure could be safely carried out.U.S. Pat. No. 4,489,053 (Azuma et al.; Dec. 18, 1984) relates toTc-99m-based diagnostic imaging agents. The patentee notes that thenon-radioactive agents may be prepared in lyophilized form and thatstabilizers are required to prevent radiolysis once the Tc-99m is added.

Thus, there is a need in the art for a method of centrally preparing andpurifying a stabilized diagnostic radiopharmaceutical for shipment tothe site of use in a form ready for simple reconstitution prior to itsadministration in diagnostic applications without the necessity ofadditional stabilizers. Because of the length of the Wolfangel process,many of the protein combinations with radionuclides are impracticalbecause of the sensitivity of the protein in combination to any freeradical attack caused by radioactive decay, and thus the presentinvention is a novel means to enable practical commercial use ofradionuclide labelled proteins and peptides. The length also effectivelyprohibits the use of shorter half life radionuclides because in order touse them with the Wolfangel process, the concentrations of theradionuclides have to be increased to account for the several half livesduring the 24 hours lyophilization and the time for shipment, whichconcentration exposes workers to higher concentrations of radioactivityand which time exposes the ligands to radiolysis which decreases theirpredictability of use in the patient, if they are effective at all. If,in order to avoid the higher concentrations, more dilute amounts areused, then the quantity of liquid involved jeopardizes the efficacy oflyophilization. There is a particular need in the art for a method ofcentrally preparing and purifying radionuclide-labeled antibodies andantibody fragments, owing to their relatively unstableimmunoreactivities once in aqueous solution. Most particularly, thisinvention enables the use of short-half-life radionuclides with ligandspotentially subject to radiolysis that are stable with useful shelf lifeat room temperatures that can be shipped in a commercially cheapermanner, and easily reconstituted.

OBJECTIVES OF THE INVENTION

An object of the invention is to accelerate the removal of water tominimize the peroxidation-related effects of radiolysis because of theaccelerated removal of water which facilitates stabilization andpredictability of concentration of a ligand or non-radioactive portionof a radiopharmaceutical because of reduced radiolysis.

An object of the invention is to use the minimization ofperoxidation-related effects to improve the preservation of the chemicalsubstituent complexes typically surrounding a radionuclide.

An object of the invention is to use small quantities at concentrationswhich enable accelerated lyophilization, longer predictable storage andovernight shipment, and increase worker safety. Corollary to thisobjective is the elimination of need for cold storage and refrigeration.

An object of the invention is to use vials with an expanded surfacearea, extremely cold temperatures and very low level pressures incombination to accelerate lyophilization.

An object of the invention is to use a two stage system to acceleratelyophilization by not only lowering vacuum pressure, but also, afterinitial removal of oxidizing agents, to extract vapor more rapidly bysupercooling gas being evacuated.

An object of the invention is to create a stable vehicle for deliveringselectively toxic radionuclides to target tissues.

DETAILED DESCRIPTION OF THE INVENTION

In contrast to the Wolfangel '556 invention which stated: “thelyophilization step itself generally takes about 24 hours to perform,”the present invention proposes to produce a stable radiopharmaceuticalcomplex by a lyophilization process which “freeze-dries” the complex infive hours or less, normally 2-4 hours, and then requires no furtherrefrigeration.

The preferred mode of the invention is utilized in conjunction withIodine-123 (“I-123” (123 being the sum of the protons and neutrons))radionuclides. The following illustrates the compositions and processesof this invention, but is not meant to limit the scope of the inventionin any way.

An I-123 labelled compound such as meta-iodo-benzyl-guanidine (“MIBG”)is prepared. The concentration is increased so that ultimately one-halfmilliliter or less will equal one dose. For example the usual does ofI-123 MIBG for a typical patient would be 10 mCi (millicuries). Becausethe half life is 12 hours, in order to allow for normal radioactivedecay in shipment so that the dose is 10 mCi upon administration, 36 mCiwould be mixed on the prior day anticipating overnight shipment.

The condensing system is heavily insulated.

A hose runs from the top or side of the stainless steel pot of theprimary condenser to the vacuum pump.

A vacuum pump capable of producing a vacuum of at least 10-4 Torr wouldbe used to evacuate the chamber. An appropriate vacuum pump is modelRV-12 available from BOCEdwards, an international company, ofWilmington, Massachusets, which can be contacted through the internet.

In order to achieve the composition contemplated in this invention, theprimary condensing coil is readied at or below −40 deg. C. Promptlyafter mixing the radiopharmaceutical composition, the vial containingthe radiopharmaceutical composition, in the preferred mode the 0.36 ml.of aqueous I-123MIBG, is stoppered with the lyophilization stopper, withthe lyophilization stopper in a position to permit passage of vapour.The vial and stopper will be fully sealed at the end of the process.

The vial(s) is (are) placed into the tray and a sufficient amount ofliquid nitrogen is poured onto the tray in order to flash freeze thevials by the heat transfer from the aqueous I-123MIBG through the sidesof the vial. Because of the small quantity which is used and the highsurface area of the vial, the freezing occurs virtually instantaneously.The tray is placed into a stoppering frame in the chamber with the innertube connected and installed so that at the end of the procedure, beforethe vacuum is broken, the port to the inner tube can be opened and thetube will inflate and force the stoppers fully into the vials in orderto seal them.

As the liquid nitrogen evaporates off, a thermistor on one of the vialsis connected to the electrical connector on the rubber stopper whichconnects to an outside temperature monitoring device. The liquidnitrogen is allowed to evaporate, all the while maintaining thetemperature of the vial at or below −10 degrees C.

The top of the chamber is installed and forms a seal with thecylindrical side of the chamber. After evaporation of the liquidnitrogen, the gas valve on top of the chamber is closed, and the rubberstopper is installed.

After the tray containing the flash-frozen vials is placed into thechamber, and the chamber has been sealed, the vacuum pump is turned on.A vacuum pressure is first felt in the primary condenser and any vaporin the chamber begins to flow out through the secondary condenser andfreezes in the primary condenser which is kept at a temperature abovethe boiling point of oxygen, meaning preferably kept at about −40degrees C. A reasonably skilled practitioner in the art would recognizethat at 10-2 Torr and −40 degree C. the amount of oxygen present wouldbe sufficiently low that the danger of oxygen oxidation damage fromliquid oxygen if the temperature is lowered below −40 degree C. iseliminated. The preferable level for activating the secondary condenseris 10-3 Torr. When the vacuum pump gauge shows the preferred level of10-3 Torr, usually after about 20 minutes, liquid Nitrogen at −196degrees C. is allowed to flow through the secondary condenser and coolthe stainless steel tube contained in the secondary condenser throughwhich gas evacuated from the chamber is flowing. The very cold liquidNitrogen in the secondary compressor is used to increase the temperaturedifference between the secondary condenser and the vial contents toaccelerate the lyophilization. The secondary condenser is placed inseries with the primary condenser and the evacuated chamber containingthe tray of vials. The secondary condenser takes over as the larger andfaster heat sink to capture the vaporized water. A reasonably skilledpractitioner would understand that the vacuum pump continues to run tothe end of the procedure, and the pressure continues to drop to therated capacity of the vacuum pump. A reasonably skilled practitionerwould know that the pump referenced, the model RV-12 available fromBOCEdwards, has a rated capacity of approximately 10-6. Thus, after thesystem has been sealed and the pump is turned on, the pressure dropsthrough the 10-2 Torr and 10-3 Torr levels to the rated capacity of thevacuum pump.

Because the acrylic chamber has no refrigeration, the temperature of thevial and the vial contents tend to rise above 0 degrees C. after all ofthe water is removed. This signals the completion of the cycle. Thethermistor probe connected through the rubber stopper to the outsidemonitoring device enables the monitoring of the vial temperature. Thevials would then be sealed in partial pressure of pharmaceutically inertgas that is fully dehydrated or “dry,” meaning gas that is non-reactivewith the pharmaceutical composition, the gas preferably being argon ornitrogen. An inner tube will have been placed in the chamber to beinflated to force the stoppers into the vial to seal them. An auxiliarycylinder of gas that is chemically inert relative to the lyophilizedradionuclide is used to gradually inflate the inner tube through thevalve to force the stoppers into the vials. The vacuum is broken. Thevial stoppers further secured with an aluminum seal. At the end of theprocess upon warming, the water which was frozen and subsequently meltedwill be drained from the primary condenser.

The vials are ready to be shipped with predictable half lives for theradionuclide and a stabilized ligand in powdered form.

If it is desired to accelerate the lyophilization process, inert gas maybe admitted through the gas valve into the chamber to displace anyoxygen and enable the secondary condenser to be turned on sooner. Thedisplacement is necessary to prevent accumulation of liquid oxygen inthe secondary condenser. In the ordinary procedure, if the secondarycondenser is activated before the 10-3 level is reached, there is a riskof collecting liquid oxygen which is potentially explosive.

The secondary condenser is in series with the primary condenser, andcould be located subsequent to the primary condenser in the evacuationand condensing system.

The speed of the lyophilization process is positively influenced by thelowering of the vapor pressure external to the material being dried.Secondly, the speed is positively influenced by the greater temperaturedifference between product being cooled and the temperature of thecondenser where the water is being collected.

The radioactive diagnostic radiopharmaceutical in this inventionrequires no further cold or refrigerated storage, including with respectto shipping, subsequent to stabilization. The lyophilizedradiopharmaceutical composition is reconstituted “on site” foradministration to patients by the addition of a suitable diluent tobring the radiopharmaceutical complex into solution at the time ofadministration to the patient.

For administration, the I-123 labelled MIBG in the vial must bereconstituted. Because of the minute quantity of material, the vial ofradionuclide complex, in the preferred mode the I-123 labelled MIBG willappear empty. The MIBG ligand is stable for several days because of theabsence of water which is the primary substance from which free radicalsare generated by gamma ray collisions with water molecules. The gammarays are being emitted by the radionuclide, that is the I-123. Thehealth care provider would add up to 2 ml. of sterile normal saline. Thedesired dose would be withdrawn and measured in a dose calibrator of atype manufactured by Capintec of Montville, N.J. If the glass vial ismeasured in the dose calibrator, the person measuring the dose mustrecognize that the glass vial will decrease the apparent activity. Uponcalibration of the desired dose, the I-123 MIBG now re-dissolved in thesolution is promptly administered to the patient.

The advantages are that the flash freezing and lowering of vaporpressure result in quick formation and evaporation or sublimation(evaporation from ice to water vapour (a gas)) of water from the I-123MIBG. The I-123 MIBG need not be shipped frozen in dry ice nor need itbe shipped for overnight delivery. Shipping in dry ice over a weekend isgenerally not commercially practical. The I-123 MIBG can be shipped overthe weekend and be used on Monday while simply maintaining it at roomtemperature or below.

In order to establish the advantages of the novel process and resultingcomposition, a series of tests were run utilizing meta iodo benzylguanidine (MIBG) in which the radionuclide I-131 was the iodine in theMIBG.

The MIBG was prepared as follows: eight vials were prepared of MIBG insolution with a radioactive concentration of MIBG of 1 mCi per vial. TheMIBG in six of those vials were then stabilized and lyophilizedaccording to the process described in this invention. One vial wasfrozen and maintained at a temperature of −10 degrees, and another vialwas simply refrigerated at approximately 5 degrees.

Six vials were prepared according to the process in this invention inorder to enable several to be reconstituted from the lyophilized stateand their activity tallied.

The radioactive concentration of MIBG per vial was 1 mCi per vial.

The results showing the percent of iodine remaining bound to the MIBGare set forth in table I. One each of the vials was reconstituted after24, 48, 72 and 168 hours respectively. 0 24 hours hours 48 hours 72hours 168 hours (1 wk.) Lyophilized 96.3% 97% 96.6%   96.2%   95.9%  and stabilized per invention stored at room temp. Frozen −10° 96.3% 94%91% 84% 72% Refrigerated 96.3% 92% 85% 77% 55% ˜+5°

In sum, the radiolysis damage was virtually eliminated from thecomposition stabilized and lyophilized under this invention while, asthe prior art suggests, MIBG that was not so stabilized and lyophilizedper this invention deteriorated sharply in activity.

As another example, I-131 Hippuran was prepared. The I-131 Hippuran wasprepared as follows: 9 vials were prepared of I-131 Hippuran in solutionwith a radioactive concentration of MIBG of 1 mCi per vial. Each vialhad 4 cc. The I-131 in seven of those vials was then stabilized andlyophilized according to the process described in this invention. Onevial was frozen and maintained at a temperature of −10 degrees, andanother vial was maintained room temperature. Room temperature wasselected because Hippuran is thought to be stable at room temperatureeven in conjunction with a radioisotope.

The results showing the percent of Hippuran remaining bound to the I-123are set forth in table 2. One each of the vials was reconstituted after24, 48, 72 and 168 hours respectively. TABLE 2 0 hours 24 hrs 48 hrs 72hrs 96 hrs 120 hrs Lyophilized 98% 98.4% 98.6%   98% 98.4% 98.5%   andstabilized per invention stored at room temp. Frozen −10° 98% 97.8% 97%94% 92.5% 91 Room Temp. 98%   96% 95% 94.5%     92% 90%

In sum, the radiolysis damage was virtually eliminated from thecomposition stabilized and lyophilized under this invention.

If one desires to ship product, maintaining a product reliably frozeneven at −10 degrees is difficult and expensive as a practical matter;this invention makes such shipment practical over the techniques of theprior art. One reference has suggested that storage at −70° C. can limitautoradiolysis damage, but even in that article, the percent freeiodine, e.g. unbonded iodine, rose from what appears to be 1.6% to 4.3%in 24 hours. Wahl, Inhibition of Autoradiolysis of Radiolabeledmonoclonal Antibodies by Cryopreservation, 31(1) J. Nucl. Med. 84-89(January 1990). Conversely, putting those results in a form analogous toTable I, the percentage of free iodine in the Wahl article commenced at98.4% and fell in 24 hours to 95.7% in Wahl's Table 1. The contrastbetween that fall in bonded iodine in 24 hours of some 3.7% in the Wahlreference versus a fall of 0.4% during a week for the compositionstabilized and lyophilized per this invention illustrates the sharpadvantage of the present method and resulting composition. In addition,it is not practical in real-world conditions to replenish the coolingfluid to maintain −70° C. much less to ship it cost-effectively.

The micro quantities involved for radionuclide complexes such as I-123MIBG substantially reduce the exposure of production workers and healthcare providers because minute quantities are involved.

More generally, the preferred mode will use compounds that have a halflife of one hour to a maximum of 12 hours. Longer half lives are lessused because of slower radioactive decay exposing the body to increasedradiation. It is generally preferable to apply the flash-freezing firstbecause application of the reduced pressure may cause the solution toboil out of the vial.

Applying the invention more generally, the intent is to utilize theinvention to produce stabilized radiopharmaceutical compositions. Suchstabilized radiopharmaceutical compositions include radionuclides whichare combined with ligand useful for diagnosis or diagnostic treatment ortherapy to form radiopharmaceutical complexes in solution or suspension.These complexes then are lyophilized in accord with the above procedureaccording to the desired radioactivity level for the selectedradionuclide. The form of radiopharmaceutical composition lyophilizedaccording to this invention can be stored until needed for use. Thisinvention allows for the central preparation, purification and shipmentof a stabilized form of a radiopharmaceutical complex which merely isreconstituted prior to use. Thus, complicated or tedious formulationprocedures, as well as unnecessary risk of exposure to radiation, at thesite of use are avoided.

The radioactive diagnostic radiopharmaceutical in this inventionrequires no further cold or refrigerated storage, including with respectto shipping, subsequent to stabilization.

The term “radiopharmaceutical composition” includes any chemicalcomposition including a radionuclide. Such term “radionuclide” includescyclotron-produced radionuclides including those referenced in Table 1on page 7 of M. Welch and C. Redvanly, Handbook of Radiopharmaceuticals:Radiochemistry and Applications (John Wiley & Sons, Ltd, Chichester,West Sussex, England 2003) (hereafter “Handbook ofRadiopharmaceuticals”), Table III on p. 77 of the Handbook ofRadiopharmaceuticals, and throughout chapters 1 and 2 of the Handbook ofRadiopharmaceuticals. Such term “radionuclide” includes reactor-producedradionuclides including those referenced in Table 2 on page 98 of theHandbook of Radiopharmaceuticals and throughout chapter 3 of theHandbook of Radiopharmaceuticals. Radionuclide also includes radioactiveisotopes of any element referenced in the Table 1 and Table 2 referencedin this paragraph, and includes Cu64 (which has traditionally not beenrecognized as useful), Fe, including Fe52 and 5959 and Fe3+radioisotopes, Yt, and Bi. Details of Gallium, Indium, and Copperradionuclides included are referenced in Tables 1 on page 264, Table 4on page 374, and Table 1 on page 402 of the Handbook ofRadiopharmaceuticals, respectively. Other useful radionuclides, whichsometimes overlap those of Table 1 and Table 2 just referenced can befound for iodine radionuclides at p. 424 of the Handbook ofRadiopharmaceuticals, and bromine radionuclides at p. 442 of theHandbook of Radiopharmaceuticals. The Technetium radionuclides andtechnetium radiopharmaceutical compositions are included. The termradiopharmaceutical composition is intended to be comprehensive becauseof the utility of the invention to radiopharmaceuticals and theirlonger-term preservation. Therefore, the term is defined to include theligands bonded with radionuclides, compounds in which the radionuclideis integral to the ligand or compound, and compounds or mixtures inwhich the radionuclide is complexed. Accordingly, further amplificationof the comprehensive scope of radiopharmaceutical composition is givenherein.

The term “radiopharmaceutical composition” includes isotopes that arebeta particle emitters, including those listed in Table 2 on page 773 ofthe Handbook of Radiopharmaceuticals, and Fe52, Cu64, Cu67, Ga68, Br77and 1124.

The term “radiopharmaceutical composition” includes radionuclides bondedto a ligand. For the purposes of this application, the term “ligand” istaken to mean a bio-compatible vehicle, typically a molecule, capable ofbinding a radionuclide and rendering the radionuclide appropriate foradministration to a patient. Thus, by way of illustration and notlimitation, the term ligand encompasses both chelating agents capable ofsequestering the radionuclide (usually a chemically-reduced form of theradionuclide) as well as carrier molecules, such as lipophilic cationswith radioisotope labeling, antibodies, antibody fragments, fatty acids,amino acids or other peptides or proteins. The term radiopharmaceuticalcomposition includes receptor specific agents, tumor agents, tumorassociated antigen, antithrombotic GPIIb/IIa receptor antagonists,agents for neuroreceptors/transporters and amyhloid plaque, BZM, andmonoclonal or polyclonal antibodies, particularly in Tcradiopharmaceuticals where preservation of the ligand is important (ageneral summary of which is on p. 349 of the Handbook ofRadiopharmaceuticals). The application of the invention to compounds forassessment of multi-drug resistance status is contemplated. Chelatingagents can include bifunctional and multifunctional chelates. Anon-exhaustive list of chelating agents is referenced on pages 366 andpage 376 of the Handbook of Radiopharmaceuticals. Included in the termligand are antibodies bound via a chelate. Such antibodies may includemonoclonal antibodies or polyclonal antibodies. Other ligandscontemplated include neuroreceptor imaging agents, and receptor imagingagents, and myocardial sympathetic nerve imaging agents, many of whichare referenced in Handbook of Radiopharmaceuticals. The carriermolecules often are specifically targeted at a tumor cell ortumor-specific antigen, an organ or a system of interest forobservational and consequent diagnostic purposes, or in need of therapy.Carrier molecules may be directly labeled with the radionuclide, inwhich case any pharmaceutically acceptable counter-ion for the therapyor diagnostic intended may be used. The radionuclide may be bound to acarrier molecule via a chelate or other binding functionality. The term“complex” is taken to mean, broadly, the union of the radionuclide andthe ligand to which it is attached. The chemical and physical nature ofthis union varies with the nature of the ligand. The invention includescompounds in the Handbook of Radiopharmaceuticals seeking receptors,including so-called antagonists which fit receptors, a partial, butfairly complete list of which is found on pages 452-457 and 717 of theHandbook of Radiopharmaceuticals.

The term “radiopharmaceutical composition” refers to a compositionincluding the radionuclide-ligand complex as well as suitablestabilizers, preservatives and/or excipients appropriate for use in thepreparation of an administrable pharmaceutical. The inventioncontemplates that for certain large proteins susceptible to breakingfrom the freezing process, such large protein structures would besupported by a lyophilization aid known to reasonably skilledpractitioners in the art of pharmacy such as lactose, dextrose, albumin,gelatin or sodium chloride.

The term “radiopharmaceutical composition”, includes, for therapeuticpurposes, therapeutic radionuclides, including Auger electron emitterssuch as those described on pages 772 and 776 of the Handbook ofRadiopharmaceuticals. Auger electron emitters can be useful because theycan result in additional deposition of energy in tissue as to whichradiopharmaceutical damage is desired. Such damage is generally desiredto be minimized in diagnostic uses.

The general method of this invention, and the composition contemplatedto be created can be implemented on a general basis as follows: after aradiopharmaceutical composition is prepared by known methods appropriateto the composition, aliquots of the radioactive complex are asepticallydispensed into sterile vials consistent with the procedure outlined andthe radioactive product is lyophilized according to the procedure ofthis invention to produce the stable lyophilized powder. The virtuallycomplete absence of water results in a substantial improvement in thestability of the preparation, from both radio chemical purity andchemical purity standpoints, versus prior preparations. The stabilizedcomplex can be prepared several days in advance, shipped and storeduntil needed for use. The preferred mode of the invention is focused onradionuclides that are gamma emitters of diagnostic value and with ahalf-life sufficiently long to make the preparation, lyophilization andshipment of the compounds practical, but the invention is useful foralpha- and beta-emitting radionuclides.

As an example of an additional preferred mode of invention, Cu64 can becomplexed with zinc isonitrile and Cu64 isonitrile can be used for PET(Positron Emission Tomography) imaging. Without the use of the processand composition of Cu64 isonitrile described herein, the half-life ofCu64 is such that its use as an imaging agent is relatively impractical.For cardiac imaging, the use of an I123 or I124 isotope in combinationwith a fatty acid is useful on a broader patient base than the currentcommonly used FDG imaging. In order to use2-deoxy-2-[18F]fluoro-D-glucose [18FDG] for imaging the heart, the heartmust be converted from fatty acid metabolism to glucose metabolism whichis accomplished by feeding the patient high levels of glucose, usuallythree or four candy bars and waiting for approximately an hour. This isunhealthy for diabetics. This invention enables the use of shorterhalf-life compounds and in particular the I123 or I-124 fatty acidradiopharmaceuticals and eliminates the necessity of conversion of theheart from fatty acid metabolism to glucose metabolism. This process andthe composition of the invention present a novel opportunity to useradioisotopes of shorter half-lives. I-124 radionuclides generally, andI-124 fatty acid radiopharmaceuticals can be used in conjunction withPET imaging.

Another preferred mode of invention is to use I124 MIBG forneuroendocrine imaging and I124 fatty acids both stabilized by thelyophilization process in this invention. Once again, only with theinvention is the use of I124 practical to sufficiently concentrate theI124 while preserving the integrity of the overall I124radiopharmaceutical composition. The use of I123 radionuclides is alsomade more practical by this invention, particularly in conjunction withfatty acid labeling.

At the point of use, the radiopharmaceutical compositions of the presentinvention are prepared for administration to a patient. Such preparationadvantageously merely involves reconstitution with an appropriatediluent to bring the complex into solution. This diluent may be sterilewater for injection (SWFI), dextrose and sodium chloride injection orsodium chloride (physiological saline) injection, for example. Thepreferred diluent is water for injection or physiological saline (9mg/ml) which conforms to the requirements listed in the U.S.Pharmacopeia.

The present invention is particularly well suited for the preparation ofstable, pre-labeled antibodies for use in the diagnosis and treatment ofcancer and other diseases. For example, antibodies expressing affinityfor specific tumors or tumor-associated antigens are labeled with adiagnostic radionuclide, either directly or via a bi-functional chelate,and the labeled antibodies are stabilized through lyophilization. Wherea bi-functional chelate is used, it generally is covalently attached tothe antibody. The antibodies used can be polyclonal or monoclonal, andthe radionuclide-labeled antibodies can be prepared according to methodsknown in the art. The method of preparation will depend upon the type ofradionuclide and antibody used. The stable, lyophilized, radio labeledantibody merely is reconstituted with suitable diluent at the time ofintended use, thus greatly simplifying the on site preparation process.The process of this invention can be applied to stabilize many types ofpre-labeled antibodies, including, but not limited to, polyclonal andmonoclonal antibodies to tumors associated with melanoma, colon cancer,breast cancer, prostate cancer, etc. Such antibodies are known in theart and are readily available. Other ligands with specific affinities tosites in need of radiotherapy are known in the art and will continue tobe discovered.

The radiopharmaceutical composition which results from the method ofthis invention may be further purified after reconstitution, if desired.One method of purification is described in EP 250966, noted above. Othermethods are known to those skilled in the art.

The radiopharmaceutical composition can include other components, ifdesired. Useful additional components include chemical stabilizers,lyophilization aids and microbial preservatives. Such chemicalstabilizers include ascorbic acid, gentisic acid, reductic acid,para-amino benzoic acid, and erythorbic acid among others. In somecases, these agents are beneficial in protecting the oxidation state ofthe radionuclide by preferential reaction with oxygen or by directeffect. The term lyophilization aids includes those substances known tofacilitate good lyophilization of the product. These aids are used toprovide bulk and stability to the dried pellet and include lactose,dextrose, albumin, gelatin, sodium chloride, mannitol, dextran andpharmaceutically-acceptable carriers, among others. Antimicrobialpreservatives inhibit the growth of or kill microbial contaminants whichare accidentally added to the product during preparation. The termantimicrobial preservatives includes methylparaben, propylparaben andsodium benzoate. These components generally are added to the compositionafter the complex has been formed between the ligand and theradionuclide but prior to lyophilization. Bacteriastatic agents, forexample, methyl and propyl-paraben may be added. Also contemplated arethe addition of solubilizing agents such as polyethylene glycol toenhance the solubility of fatty acid compounds tagged with radionuclidesin normal saline solution or other water based solutions.

The above process, apparatus and resulting composition is adaptable tothe stabilization and preservation of virtually all radionuclideswhatever the solvent used for initial composition. Some preferredapplications include stabilization of radiolabeled peptides, [18 F]deoxyglucose, radiolabelled annexin, 99 mTc-annexin, radiolabelledmonocyte chemoattractant protein. i.e. 125-I-(MCP-1), radiolabelledDopamine transporter agents,(S)-N-(1-ethylpyrrolidin-2-ylmethyl)-2-hydroxy-3-iodo-6-methoxybenzamide(3-IBZM)(More generally “BZM,),(S)-N-(1-ethylpyrrolidin-2-ylmethyl)-2-hydroxy-5-iodo-6-methoxybenzamide(5-IBZM), 1-123-2-beta-carbomethoxy-3-beta(4-iodophenyl) N-(3-fluropropyl) nortropane (“CIT” or “beta-CIT”) and various tropanederivatives, I-123 fatty acids, particularly for cardiovascular imaging,radiolabelled octreotide or radiolabelled depreotide, HEDP (diagnosticskeletal imaging or treatment of metastatic bone pain), radiolabelledantibodies, both polyclonal and monoclonal, with selective affinitiesfor tumor-associated antigens diagnosis or in situ radiotherapy ofmalignant tumors such as melanomas), and ligands with selective affinityfor the hepatobiliary system (the liver-kidney system), including2,6-dimethylacetanilideiminodiacetic acid and the family of otherimidoacetic acid group-containing analogs thereof (collectively referredto herein as “HIDA agents”), mono-, di- and polyphosphoric acids andtheir pharmaceutically-acceptable salts including polyphosphates,pyrophosphates, phosphonates, diphosphonates and imidophosphonates.Preferred ligands are 1-hydroxyethylidene diphosphonate, methylenediphosphonate, (dimethylamino)methyl diphosphonate,methanehydroxydiphosphonate, and imidodiphosphonate (for bone-scanningand alleviation of pain); strontium 89 ethylene diamine tetramethylenephosphate, samarium 153-ethylene diamine tetramethylene phosphate,radiolabelled monoclonal antibodies, 99m-Tc HMPAO (hexamethylproplyeneamine oxime), yttrium 90-labeled ibritumomab tiuxetan (Zevalin®Registered Trademark of Biogen Idec, Inc.), and meta-iodo-benzylguanidine. Ethylene diamine tetramethylene phosphate and ethylenediamine tetramethylene phosphoric acid and the pharmaceutically relatedmono-, di- and polyphosphoric acids and theirpharmaceutically-acceptable salts including polyphosphates,pyrophosphates, phosphonates, diphosphonates and imidophosphonates arecollectively called EDTMP.

Suitable radionuclides which are well-known to those skilled in the artinclude radioisotopes of copper, technetium-99m, rhenium-186,rhenium-188, antimony-127, lutetium-177, lanthanum-140, samarium-153,radioisotopes of iodine, indium-111, gallium-67 and -68, chromium-51,strontium-89, radon-222, radium-224, actinium-225, californium-246 andbismuth-210. Other suitable radionuclides include F-18, C-11, Y-90,Co-55, Zn-62, Fe-52, Br-77, Sr-89, Zr-89, Sm-153, Ho-166, and Tl-201.

The invention is not meant to be limited to the disclosures, includingbest mode of invention herein, and contemplates all equivalents to theinvention and similar embodiments to the invention for humans, mammalsand plant science. Equivalents include combinations with or withoutstabilizing agents and adjuncts that assist in reservation, and theirpharmacologically active racemic mixtures, diastereomers and enantiomersand their pharmacologically acceptable salts in combination withsuitable pharmaceutical carriers.

1. A method of preparing a stable rapidly lyophilizedradiopharmaceutical composition for diagnostic or therapeutic purposesthat needs no refrigeration upon completion of the method and thatincreases the predictability of the integrity of the radiopharmaceuticalcomposition by reducing radiolysis damage, comprising the followingsteps: evacuating a sealable chamber containing a flash frozen amount ofsaid radiopharmaceutical composition having at least one radionuclide inat least one lyophilization-stoppered but as yet unsealed vial, saidevacuating of said sealable chamber occurring by a vacuum pump connectedby an evacuation tube passing through a primary condenser and asecondary condenser to lower pressure to below 10-2 Torr which issufficient to eliminate the explosive potential of liquid oxygen whilemaintaining the temperature of said primary condenser above the boilingpoint of oxygen at said pressure; activating said secondary condenser toreduce said evacuation tube temperature below the boiling point ofoxygen in order to accelerate the removal of water from said sealablechamber, thereby reducing more rapidly the presence of water molecules,including radiolysis degenerated water molecules, and reducing attendantfree radical damage to said radiopharmaceutical composition, andincreasing the predictability of the integrity of theradiopharmaceutical composition; and restoring the ambient pressure inthe sealable chamber to approximately atmospheric pressure with apharmaceutically inert gas upon completion of the desired removal ofwater; and sealing the said at least one vial in order to preclude entryof external fluid.
 2. The method according to claim 1, furthercomprising: said evacuating said sealable chamber occurring at a primarycondenser temperature of approximately −40 degrees C. until saidpressure sufficient to eliminate the explosive potential of liquidoxygen has reached approximately 10-2 Torr.
 3. The method according toclaim 2, further comprising: said radiopharmaceutical composition havingat least one ligand.
 4. The method according to claim 3, furthercomprising: said at least one ligand being selected from the group ofBZM, Beta-CIT, EDTMP, HIDA or fatty acids.
 5. The method according toclaim 3, further comprising: said radiopharmaceutical composition havingthe ligand MIBG.
 6. The method according to claim 2, further comprising:said radiopharmaceutical composition having at least one monoclonalantibody in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemonoclonal antibody.
 7. The method according to claim 2, furthercomprising: said radiopharmaceutical composition having at least onepeptide in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onepeptide.
 8. The method according to claim 2, further comprising: saidradiopharmaceutical composition having at least one molecularrecognition unit in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemolecular recognition unit.
 9. The method according to claim 2, furthercomprising: said at least one radionuclide being selected from the groupof F-18, C-11, Y-90, I-123, I-124, I-125, I-131, Cu-64, Cu-67, Co-55,Zn-62, Fe-52, Ga-64, Ga-67, Ga-68, Br-77, Sr-89, Zr-89, Tc-99m, In-111,Sm-153, Ho-166, Lu-177, Re-186, and Tl-201.
 10. The method according toclaim 9, further comprising: said radiopharmaceutical composition havingat least one ligand.
 11. The method according to claim 10, furthercomprising: said at least one ligand being selected from the group ofBZM, Beta-CIT, EDTMP, HIDA or fatty acids.
 12. The method according toclaim 10, further comprising: said radiopharmaceutical compositionhaving the ligand MIBG.
 13. The method according to claim 9, furthercomprising: said radiopharmaceutical composition having at least onemonoclonal antibody in combination with at least one lyophilization aidfor providing structural stabilization in combination with said at leastone monoclonal antibody.
 14. The method according to claim 9, furthercomprising: said radiopharmaceutical composition having at least onepeptide in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onepeptide.
 15. The method according to claim 9, further comprising: saidradiopharmaceutical composition having at least one molecularrecognition unit in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemolecular recognition unit.
 16. The method according to claim 1, furthercomprising: said radiopharmaceutical composition having at least oneligand.
 17. The method according to claim 16, further comprising: saidat least one ligand being selected from the group of BZM, Beta-CIT,EDTMP, HIDA or fatty acids.
 18. The method according to claim 16,further comprising: said radiopharmaceutical composition having theligand MIBG.
 19. The method according to claim 1, further comprising:said radiopharmaceutical composition having at least one monoclonalantibody in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemonoclonal antibody.
 20. The method according to claim 1, furthercomprising: said radiopharmaceutical composition having at least onepeptide in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onepeptide.
 21. The method according to claim 1, further comprising: saidradiopharmaceutical composition having at least one molecularrecognition unit in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemolecular recognition unit.
 22. The method according to claim 1, furthercomprising: said at least one radionuclide being selected from the groupof F-18, C-11, Y-90, I-123, I-124, I-125, I-131, Cu-64, Cu-67, Fe-52,Co-55, Zn-62, Ga-64, Ga-67, Ga-68, Br-77, Sr-89, Zr-89, Tc-99m, In-111,Sm-153, Ho-166, Lu-177, Re-186, and Tl-201.
 23. The method according toclaim 22, further comprising: said radiopharmaceutical compositionhaving at least one ligand.
 24. The method according to claim 23,further comprising: said at least one ligand being selected from thegroup of BZM, Beta-CIT, EDTMP, HIDA or fatty acids.
 25. The methodaccording to claim 23, further comprising: said radiopharmaceuticalcomposition having the ligand MIBG.
 26. The method according to claim22, further comprising: said radiopharmaceutical composition having atleast one monoclonal antibody in combination with at least onelyophilization aid for providing structural stabilization in combinationwith said at least one monoclonal antibody.
 27. The method according toclaim 22, further comprising: said radiopharmaceutical compositionhaving at least one peptide in combination with at least onelyophilization aid for providing structural stabilization in combinationwith said at least one peptide.
 28. The method according to claim 22,further comprising: said radiopharmaceutical composition having at leastone molecular recognition unit in combination with at least onelyophilization aid for providing structural stabilization in combinationwith said at least one molecular recognition unit.
 29. A method ofpreparing a stable rapidly lyophilized radiopharmaceutical compositionfor diagnostic or therapeutic purposes that needs no refrigeration uponcompletion of the method and that increases the predictability of theintegrity of the radiopharmaceutical composition by reducing radiolysisdamage, comprising the following steps: evacuating a sealable chambercontaining a flash frozen amount of said radiopharmaceutical compositionhaving at least one radionuclide in at least onelyophilization-stoppered but as yet unsealed vial, said said evacuatingof said sealable chamber occurring by a vacuum pump through anevacuation tube passing through a secondary condenser to a primarycondenser to lower pressure to below 10-2 Torr which is sufficient toeliminate the explosive potential of liquid oxygen while maintaining thetemperature of said primary condenser for cooling above the boilingpoint of oxygen at said pressure; activating said secondary condenser toreduce said evacuation tube temperature below the boiling point ofoxygen in order to accelerate the removal of water from said sealablechamber, thereby reducing more rapidly the presence of water molecules,including radiolysis degenerated water molecules, and reducing attendantfree radical damage to said radiopharmaceutical composition, andincreasing the predictability of the integrity of theradiopharmaceutical composition; and restoring the ambient pressure inthe sealable chamber to approximately atmospheric pressure with apharmaceutically inert gas upon completion of the desired removal ofwater; and sealing said at least one vial in order to preclude entry ofexternal fluid.
 30. The method according to claim 29, furthercomprising: said evacuating said sealable chamber occurring at a primarycondenser temperature of approximately −40 degrees C. until saidpressure sufficient to eliminate the explosive potential of liquidoxygen has reached approximately 10-2 Torr.
 31. The method according toclaim 30, further comprising: said radiopharmaceutical compositionhaving at least one ligand.
 32. The method according to claim 31,further comprising: said at least one ligand being selected from thegroup of BZM, Beta-CIT, EDTMP, HIDA or fatty acids.
 33. The methodaccording to claim 31, further comprising: said radiopharmaceuticalcomposition having the ligand MIBG.
 34. The method according to claim30, further comprising: said radiopharmaceutical composition having atleast one monoclonal antibody in combination with at least onelyophilization aid for providing structural stabilization in combinationwith said at least one monoclonal antibody.
 35. The method according toclaim 30, further comprising: said radiopharmaceutical compositionhaving at least one peptide in combination with at least onelyophilization aid for providing structural stabilization in combinationwith said at least one peptide.
 36. The method according to claim 30,further comprising: said radiopharmaceutical composition having at leastone molecular recognition unit in combination with at least onelyophilization aid for providing structural stabilization in combinationwith said at least one molecular recognition unit.
 37. The methodaccording to claim 30, further comprising: said at least oneradionuclide being selected from the group of F-18, C-11, Y-90, I-123,I-124, I-125, I-131, Cu-64, Cu-67, Co-55, Zn-62, Fe-52, Ga-64, Ga-67,Ga-68, Br-77, Sr-89, Zr-89, Tc-99m, In-111, Sm-153, Ho-166, Lu-177,Re-186, and Tl-201.
 38. The method according to claim 37, furthercomprising: said radiopharmaceutical composition having at least oneligand.
 39. The method according to claim 38, further comprising: saidat least one ligand being selected from the group of BZM, Beta-CIT,EDTMP, HIDA or fatty acids.
 40. The method according to claim 38,further comprising: said radiopharmaceutical composition having theligand MIBG.
 41. The method according to claim 37, further comprising:said radiopharmaceutical composition having at least one monoclonalantibody in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemonoclonal antibody.
 42. The method according to claim 37, furthercomprising: said radiopharmaceutical composition having at least onepeptide in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onepeptide.
 43. The method according to claim 37, further comprising: saidradiopharmaceutical composition having at least one molecularrecognition unit in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemolecular recognition unit.
 44. The method according to claim 29,further comprising: said radiopharmaceutical composition having at leastone ligand.
 45. The method according to claim 44, further comprising:said at least one ligand being selected from the group of BZM, Beta-CIT,EDTMP, HIDA or fatty acids.
 46. The method according to claim 44,further comprising: said radiopharmaceutical composition having theligand MIBG.
 47. The method according to claim 29, further comprising:said radiopharmaceutical composition having at least one monoclonalantibody in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemonoclonal antibody.
 48. The method according to claim 29, furthercomprising: said radiopharmaceutical composition having at least onepeptide in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onepeptide.
 49. The method according to claim 29, further comprising: saidradiopharmaceutical composition having at least one molecularrecognition unit in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemolecular recognition unit.
 50. The method according to claim 29,further comprising: said at least one radionuclide being selected fromthe group of F-18, C-11, Y-90, I-123, I-124, I-125, I-131, Cu-64, Cu-67,Fe-52, Co-55, Zn-62, Ga-64, Ga-67, Ga-68, Br-77, Sr-89, Zr-89, Tc-99m,In-111, Sm-153, Ho-166, Lu-177, Re-186, and Tl-201.
 51. The methodaccording to claim 50, further comprising: said radiopharmaceuticalcomposition having at least one ligand.
 52. The method according toclaim 51, further comprising: said at least one ligand being selectedfrom the group of BZM, Beta-CIT, EDTMP, HIDA or fatty acids.
 53. Themethod according to claim 51, further comprising: saidradiopharmaceutical composition having the ligand MIBG.
 54. The methodaccording to claim 50, further comprising: said radiopharmaceuticalcomposition having at least one monoclonal antibody in combination withat least one lyophilization aid for providing structural stabilizationin combination with said at least one monoclonal antibody.
 55. Themethod according to claim 50, further comprising: saidradiopharmaceutical composition having at least one peptide incombination with at least one lyophilization aid for providingstructural stabilization in combination with said at least one peptide.56. The method according to claim 50, further comprising: saidradiopharmaceutical composition having at least one molecularrecognition unit in combination with at least one lyophilization aid forproviding structural stabilization in combination with said at least onemolecular recognition unit.
 57. The method according to claims 1 through56, further comprising: said radiopharmaceutical composition being animaging agent selected from the group of imaging agents having aselective affinity for the hepatobiliary system.
 58. The methodaccording to claims 1 through 56, further comprising: saidradiopharmaceutical composition being an imaging agent selected from thegroup of imaging agents having a selective affinity for the cardiacsystem.
 59. The method according to claims 1 through 56, furthercomprising: said radiopharmaceutical composition being an imaging agentselected from the group of imaging agents having a selective affinityfor the cerebral system.
 60. The method according to claims 1 through56, further comprising: said radiopharmaceutical composition being animaging agent selected from the group of imaging agents having aselective affinity for the skeletal system.
 61. The method according toclaims 1 through 56, further comprising: said radiopharmaceuticalcomposition being an imaging agent selected from the group of imagingagents used for prostate imaging.
 62. The method according to claims 1through 56, further comprising: said radiopharmaceutical compositionbeing an imaging agent selected from the group of imaging agents usedfor pulmonary imaging.
 63. The method according to claims 1 through 56,further comprising: said radiopharmaceutical composition having at leastone chemical stabilizer.
 64. The method according to claims 1 through56, further comprising: said radiopharmaceutical composition having atleast one bacteriastatic agent.
 65. The method according to claims 1through 56, further comprising: said radiopharmaceutical compositionhaving at least one antimicrobial preservative.
 66. The method accordingto claims 1 through 56, further comprising: said radiopharmaceuticalcomposition having at least one solubilizing agent.
 67. The methodaccording to claims 1-5, 9-12, 16-18, 22-25, 29-33, 37-40, 44-46, and50-53, further comprising: said radiopharmaceutical compositioncomprising at least one lyophilization aid.
 68. The method according toclaims 1-5, 9-12, 16-18, 22-25, 29-33, 37-40, 44-46, and 50-53, furthercomprising: said radiopharmaceutical composition comprising at least onelyophilization aid selected from the group of lactose, dextrose,albumin, gelatin or sodium chloride.
 69. The method according to claims6, 13, 19, 26, 34, 41, 47, and 54, further comprising: saidradiopharmaceutical composition having at least one monoclonal antibodyin combination with at least one lyophilization aid selected from thegroup of lactose, dextrose, albumin, gelatin or sodium chloride forproviding structural stabilization in combination with said at least onemonoclonal antibody.
 70. The method according to claims 7, 14, 20, 27,35, 42, 48, and 55, further comprising: said radiopharmaceuticalcomposition having at least one peptide in combination with at least onelyophilization aid selected from the group of lactose, dextrose,albumin, gelatin or sodium chloride for providing structuralstabilization in combination with said at least one peptide.
 71. Themethod according to claims 8, 15, 21, 28, 36, 43, 49, and 56, furthercomprising: said radiopharmaceutical composition having at least onemolecular recognition unit in combination with at least onelyophilization aid selected from the group of lactose, dextrose,albumin, gelatin or sodium chloride for providing structuralstabilization in combination with said at least one molecularrecognition unit.